Several competing models have been proposed to explain how neurofilaments (NFs), the most abundant intermediate filament system in neurons, interact to form a structural framework that mechanically stabilizes the axon. In one recently developed model, adjacent NFs interact through mutual steric repulsion, whose range and strength is modulated by the degree of NF phosphorylation. I propose to quantitatively test this hypothesis in living cells through two specific aims. First, to explore the contribution of NF phosphorylation to whole-cell mechanical properties, I will perturb the NF CSK pharmacologically, biochemically, and mechanically and measure changes in mechanical properties using traction force microscopy, magnetic micromanipulation techniques, and confocal fluorescence microscopy. Second, using these same perturbations, I will determine the effect of NF integrity and phosphorylation to two cell-shape dependent properties, cell fate determination and neurite extension. By clarifying the macromolecular origins of neuronal shape stability, these studies will improve our insight into the role of cell shape changes in neuronal pathology and may suggest novel modes of therapeutic intervention.